This application addresses broad Challenge Area (15) Translational Science and specific Challenge Topic, 15-DK-101- Identification of bioactive macronutrients in the diet that impact metabolic state. Type 2 Diabetes is epidemic in western populations. Insulin resistance, stemming in part from a state of "oxidative stress" centered in mitochondria, is an early event that both predicts and contributes to the onset of diabetes. While several classes of drugs are currently prescribed to enhance insulin sensitivity, these therapies are hindered by an increased propensity for adverse cardiovascular events (including heart failure and infarction). Diets rich in leafy green vegetables and fish, such as the Mediterranean diet, are associated with a striking reduction in obesity, diabetes and overall cardiovascular risk. Although the "active ingredient" in this diet remains uncertain, recent studies demonstrate that the simple salt nitrate (NO3-), upon metabolism to nitrite (NO2-), supports the generation of signaling mediators. Preliminary data show that the mitochondrion acts as a "bioreactor" to catalyze the modification of proteins and lipids by NO2-, yielding salutary cell signaling species including S-nitrosated proteins and nitrated lipids, which regulate fundamental physiological events. Specifically, these species can regulate mitochondrial function and the expression of inflammatory-related genes via post-translational protein modification and activation of the nuclear lipid receptor PPAR3. Here it is hypothesized that dietary nitrate is a bioactive dietary nutrient that is converted to species that modulate oxidative inflammatory reactions and metabolism and activates PPAR3, thereby increasing insulin sensitivity. A strong team of basic and clinical physiologists and biochemists with expertise in redox-dependent cell signaling, cardiovascular biology and diabetes has devised a readily executable research plan to test this hypothesis. The effect of dietary nitrate on insulin sensitivity will be tested using a novel technique of frequently sampled intravenous glucose tolerance tests and hyperinsulinemic euglycemic clamps in chronically catheterized wild type and obese mice. Detailed mechanistic understanding will be obtained, using a targeted biochemical approach, regarding how dietary NO3- and NO2- can influence insulin secretion and sensitivity via mitochondrial redox reactions. The knowledge gained from this study will initiate a new understanding of mechanisms underlying insulin sensitivity and a novel pharmacologic strategy for treating T2D. Characterization of the signaling reactions of this dietary molecule abundant in "heart-healthy" foods will improve the general understanding of how simple lifestyle changes can yield profound benefit for managing and preventing insulin sensitivity, thereby increasing the likelihood that members of the general population will commit to these sustained lifestyle changes. Type 2 Diabetes is epidemic in westernized populations and insulin resistance contributes to the onset of diabetes. PUBLIC HEALTH RELEVANCE: A molecule called nitrate, found in leafy green vegetables and other components of a healthy diet, may play an important role in preventing and reversing insulin resistance. Understanding how nitrate works with regards to diabetes will help researchers and physicians develop scientifically-based dietary guidelines for people with diabetes.